A vaccine for prevention of genital herpes is a high public health priority. We are evaluating a trivalent subunit antigen vaccine that contains herpes simplex virus type 2 (HSV-2) glycoproteins C, D, and E (gC2, gD2, gE2). Glycoprotein D is involved in virus entry and cell-to-cell spread, while gC2 and gE2 are immune evasin molecules that inhibit complement and antibody. We used the Carterra high throughput biosensor technology to evaluate whether immunization with gD2 subunit antigen in guinea pigs produced antibodies to each of seven crucial gD2 epitopes involved in virus attachment to cells, activation of downstream entry molecules glycoproteins H and L (gH2/gL2), or cell-to-cell spread. Unexpectedly, we found that gD2-immunized guinea pigs produced antibodies that blocked on average only four of seven crucial epitopes. Some animals blocked 2 epitopes, while others blocked six or seven. The more epitopes blocked, the better was the protection against intravaginal HSV-2 challenge. We compared the epitope-specific results in guinea pigs with our earlier studies of subjects enrolled in the GlaxoSmithKline gD2 Herpevac Trial for Women. Humans responded to even fewer epitopes than guinea pigs. Some epitopes were weak immunogens in humans and guinea pigs, including one that mediates receptor attachment and another involved in cell-to-cell spread. Our goal in this Phase 1 study is to improve the immunogenicity of weakly immunogenic epitopes. We will use EpiSweep computer-based structural technology to stealth or de-immunize epitopes that are highly immunogenic in an effort to enhance the immunogenicity of weakly immunogenic epitopes. We will focus initially on stealthing a gD2 epitope that interacts with gH2/gL2 during entry. This epitope is highly immunogenic in guinea pigs and humans. We will combine a computer-based approach with our knowledge of gD2 structure to develop gD2 constructs with altered amino acid sequences. We will prepare these constructs as subunit antigens in baculovirus and use the Carterra biosensor platform and our large panel of gD2 monoclonal antibodies to determine whether we have stealthed the epitope as intended. We will then immunize guinea pigs with the modified antigens and use the Carterra platform to determine whether antibodies are produced to the previously weakly immunogenic epitopes. If the stealthing process results in an unintended loss of immunogenicity of the strong epitope, we will boost with unmodified gD2 subunit antigen to stimulate a response. Our focus in this Phase 1 study is to improve the immunogenicity of gD2. Our longer-term goal is to use this technology to improve epitope responses to gC2 and gE2 antigens in the vaccine.